The three enzymes of 2,3-diphosphoglycerate metabolism are: monophosphoglycerate mutase, diphosphoglycerate mutase, and 2,3- diphosphoglycerate phosphatase. The first is a glycolytic enzyme and the other two are responsible for the synthesis and breakdown of 2,3- diphosphoglycerate (2,3-DPG). The latter is important in human red blood cells since it has been found that the presence of 2,3-DPG decreases the oxygen affinity of hemoglobin so that oxygen is released efficiently to the tissues. We have purified the latter two enzymes from human red blood cells and are studying their properties and the regulation of the level of 2,3-DPG in the cell. Previous studies of the enzyme have shown that both monophosphoglycerate mutase and diphosphoglycerate mutase can be phosphorylated by 2,3-DPG. The role of phosphorylation in the catalytic reactions is to be determined. Kinetic studies of the phosphatase have been carried out. We are trying to prepare larger amounts of phosphatase, free of the other two enzymes, to study whether phosphorylation occurs and to characterize its molecular properties, i.e. molecular weight and subunit structure. The possible evolutionary inter-relationship of the enzymes will be considered.